[Anti-asthma components and mechanism of Kechuanting acupoint application therapy: based on serum metabolomics and network pharmacology].

This study aims to explore the anti-asthma components and mechanism of Kechuanting acupoint application therapy(KAAT) based on serum metabolomics and network pharmacology. A total of 60 asthma patients who had used low-dose inhaled corticosteroids-formoterol(ICS-formoterol) for a long time were randomized into the western medicine group(low-dose ICS-formoterol) and western medicine+Kechuanting group(KAAT+low-dose ICS-Formoterol), 30 in either group. In addition, 30 healthy people were included as the control(no intervention). The asthma control test(ACT) score, forced expiratory volume in 1 second(FEV1), and peak expiratory flow(PEF) were measured in the western medicine group and western medicine+Kechuanting group before and after treatment. The potential biomarkers of KAAT in the treatment of asthma were screened by gas chromatography-mass spectrometry combined with multivariate analysis, and the related metabolic pathways were further analyzed. UPLC/LTQ-Orbitrap-MS, together with network pharmacology, was employed to construct the component-target-pathway network. Thereby, the effective components and me-chanism of KAAT in the treatment of asthma were clarified. According to the ACT score, FEV1, and PEF, KAAT was effective in the treatment of asthma. A total of 10 endogenous biomarkers of KAAT in the treatment of asthma were screened by serum metabolomics, and the pathways of the metabolism of glycine, serine and threonine, and the metabolism of glyoxylic acid and dicarboxylic acid were obtained. UPLC/LTQ-Orbitrap-MS identified 51 chemical components of KAAT: 24 flavonoids, 11 alkaloids, 8 phenols, 2 diterpenoids, 2 triterpenoids, 2 glycosides, and 2 aldehydes. Network pharmacology analysis suggested that KAAT mainly acted on serum crea-tinine(SRC), matrix metalloproteinase 9(MMP-9), and other target proteins. The treatment was closely related to metabolic pathway, phosphatidylinositol 3-kinase-protein kinase B(PI3 K-Akt), mitogen-activated protein kinase(MAPK), and calcium signaling pathway. Sinapine thiocyanate, corydaline, dihydroberberine, stylopine, leonticine, N-methyl tetrahydroberberine, kaempferide, erio-dictyol, quercetin, catechin, 6-gingerol, 6-shogaol, ingenol, and luteolin may be potential effective compounds of KAAT in the treatment of asthma. This study preliminarily revealed that the effective components and mechanism of KAAT in treatment of asthma based on serum metabolomics and network pharmacology. It lays a theoretical foundation for in-depth study of the mechanism and clinical development and application.

[Components of drugs in acupoint sticking therapy and its mechanism of intervention on bronchial asthma based on UPLC-Q-TOF-MS combined with network pharmacology and experimental verification].

UPLC-Q-TOF-MS combined with network pharmacology and experimental verification was used to explore the mechanism of acupoint sticking therapy(AST) in the intervention of bronchial asthma(BA). The chemical components of Sinapis Semen, Cory-dalis Rhizoma, Kansui Radix, Asari Radix et Rhizoma, and Zingiberis Rhizoma Recens were retrieved from TCMSP as self-built database. The active components in AST drugs were analyzed by UPLC-Q-TOF-MS, and the targets were screened out in TCMSP and Swiss-TargetPrediction. Targets of BA were collected from GeneCards, and the intersection of active components and targets was obtained by Venny 2.1.0. The potential targets were imported into STRING and DAVID for PPI, GO, and KEGG analyses. The asthma model induced by house dust mite(HDM) was established in mice. The mechanism of AST on asthmatic mice was explored by pulmonary function, Western blot, and flow cytometry. The results indicated that 54 active components were obtained by UPLC-Q-TOF-MS and 162 potential targets were obtained from the intersection. The first 53 targets were selected as key targets. PPI, GO, and KEGG analyses showed that AST presumedly acted on SRC, PIK3 CA, and other targets through active components such as sinoacutine, sinapic acid, dihydrocapsaicin, and 6-gingerol and regulated PI3 K-AKT, ErbB, chemokine, sphingolipid, and other signaling pathways to intervene in the pathological mechanism of BA. AST can improve lung function, down-regulate the expression of PI3 K and p-AKT proteins in lung tissues, enhance the expression of PETN protein, and reduce the level of type Ⅱ innate immune cells(ILC2 s) in lung tissues of asthmatic mice. In conclusion, AST may inhibit ILC2 s by down-regulating the PI3 K-AKT pathway to relieve asthmatic airway inflammation and reduce airway hyperresponsiveness.

Improvement of long-term clinical outcomes by successful PCI in the very elderly women with ACS.

BACKGROUND: Whether very elderly women with acute coronary syndromes (ACS) should receive aggressive percutaneous coronary intervention (PCI) is still controversial. We assessed the effectiveness and long-term clinical outcomes of successful PCI in this population and identified prognostic factors which might contribute to the incidence of major adverse cardiovascular and cerebrovascular events (MACCE) in the very elderly female PCI cohort. METHODS: Female ACS patients aged ≥ 80 years were consecutively enrolled (n = 729) into the study. All the patients were divided into female PCI group (n = 232) and medical group (n = 497). MACCE was followed up, including non-fatal myocardial infarction (MI), stroke, heart failure requiring hospitalization (HFRH), cardiovascular (CV) death, and the composite of them. After propensity score matching (1:1), the incidences of MACCE were compared between the two groups. Clinical and coronary artery lesion characteristics were compared between the female PCI patients with (n = 56) and without MACCE (n = 176). Multivariate Cox regression analysis was performed to identify risk factors which independently associated with MACCE in the female PCI patients. MACCE of male PCI patients, who aged ≥ 80 years and hospitalized in the same period (n = 264), was also compared with that of the female PCI patients. RESULTS: A total of 32% very elderly female ACS patients received PCI in the present study. (1) Compared to female medical group, PCI procedure significantly alleviated the risks of MACCE: non-fatal MI (6.2% vs. 20.2%, P < 0.001), HFRH (10.9% vs. 22.5%, P = 0.012), CV death (12.4% vs. 28.7%, P < 0.001) and the composite MACCE (24.0% vs. 44.2%, P < 0.001) during the median follow-up period of 36 months. (2) Between very elderly female and male PCI patients, there were no significant differences in occurrence of MACCE (P = 0.232) and CV death (P = 0.951). (3) Multivariate Cox analysis revealed that ST-segment elevation myocardial infarction (STEMI) (HR 1.944, 95% CI 1.11-3.403, P = 0.02) and elevated log- N-Terminal pro-brain natriuretic peptide (NT-proBNP) (HR 1.689, 95% CI 1.029-2.773, P = 0.038) were independently associated with the incidence of MACCE in the female PCI patients. CONCLUSIONS: PCI procedure significantly attenuated the risk of MACCE and improved the long-term clinical outcomes in very elderly female ACS patients. Aggressive PCI strategy may be reasonable in this population.

Attenuation of High Glucose-Induced Rat Cardiomyocyte Apoptosis by Exendin-4 via Intervention of HO-1/Nrf-2 and the PI3K/AKT Signaling Pathway.

Exendin-4, a glucagon-like peptide-1 receptor agonist, demonstrated cytoprotective actions beyond glycemic control in recent studies. The aims of the present study were to investigate the effects of exendin-4 on high glucose (HG)-induced cardiomyocyte apoptosis and the possible mechanisms. Rat cardiomyocytes were divided into 3 groups: normal glucose group (NG group), HG group and HG +exendin-4 group (HG+Ex Group). Cardiomyocyte apoptosis was evaluated by double-staining with annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) and flow cytometry. Intracellular reactive oxygen species (ROS) production was detected by 2',7'-dichlorodihydrofluorescein diacetate (DCHF-DA) incubation and fluorescence microscopy. LY294002 (LY), a phosphoinositide 3-kinase (PI3K) pathway inhibitor, was added to the medium of the HG+Ex+LY Group for further western blot analysis. The proteins analyzed involved oxidative stress-associated proteins, heme oxygenase-1 (HO-1) and nuclear factor E2-related factor 2 (Nrf-2), and apoptosis-associated proteins, caspase-3, Bax/B-cell lymphoma 2 (Bcl-2) and p-AKT/AKT. HG treatment induced cardiomyocyte apoptosis (P = 0.00) and clearly upregulated ROS production (P = 0.00); exendin-4 co-incubation also ameliorated cardiomyocyte apoptosis (P = 0.004) and decreased ROS (P = 0.00) level significantly. HO-1 and Nrf-2 protein expression levels decreased significantly in the HG group (P < 0.05), but the levels were elevated by exendin-4 intervention (P < 0.05). Furthermore, exendin-4 attenuated HG-induced higher protein expression, including cleaved caspase-3 and Bax, increased the expression of Bcl-2 protein (P < 0.05). However, these impacts of exendin-4 were counteracted significantly by co-incubation with LY294002. In addition, exendin-4 ameliorated HG-induced p-AKT/AKT lower expression, and this impact was also suppressed by LY294002. Exendin-4 ameliorates HG-induced cardiomyocyte apoptosis, and the mechanisms may involve anti-oxidative stress via the HO-1/Nrf-2 system, as well as intervention of the PI3K/AKT signaling pathway.

Progressive decay of Ca2+ homeostasis in the development of diabetic cardiomyopathy.

BACKGROUND: Cardiac dysfunction in diabetic cardiomyopathy may be associated with abnormal Ca2+ homeostasis. This study investigated the effects of alterations in Ca2+ homeostasis and sarcoplasmic reticulum Ca2+-associated proteins on cardiac function in the development of diabetic cardiomyopathy. METHODS: Sprague-Dawley rats were divided into 4 groups (n = 12, each): a control group, and streptozotocin-induced rat models of diabetes groups, examined after 4, 8, or 12 weeks. Evaluations on cardiac structure and function were performed by echocardiography and hemodynamic examinations, respectively. Cardiomyocytes were isolated and spontaneous Ca2+ spark images were formed by introducing fluorescent dye Fluo-4 and obtained with confocal scanning microscopy. Expressions of Ca2+-associated proteins were assessed by Western blotting. RESULTS: Echocardiography and hemodynamic measurements revealed that cardiac dysfunction is associated with the progression of diabetes, which also correlated with a gradual but significant decline in Ca2+ spark frequency (in the 4-, 8- and 12-week diabetic groups). However, Ca2+ spark decay time constants increased significantly, relative to the control group. Expressions of ryanodine receptor 2 (RyR2), sarcoplasmic reticulum Ca2+-2ATPase (SERCA) and Na+/Ca2+ exchanger (NCX1) were decreased, together with quantitative alterations in Ca2+regulatory proteins, FKBP12.6 and phospholamban progressively and respectively in the diabetic rats. CONCLUSIONS: Ca2+ sparks exhibited a time-dependent decay with progression of diabetic cardiomyopathy, which may partly contribute to cardiac dysfunction. This abnormality may be attributable to alterations in the expressions of some Ca2+-associated proteins.

Prognostic Value of NT-proBNP in Patients With Successful PCI for ACS and Normal Left Ventricular Ejection Fraction.

BACKGROUND: Patients undergoing successful percutaneous coronary intervention (PCI) for acute coronary syndromes (ACS) with normal left ventricular ejection fraction (LVEF) are generally considered to have successful clinical outcomes; however, there are still significant differences in clinical outcomes among these patients. The aim of the study was to find a common indicator to predict the risk of major adverse cardiac and cerebrovascular events (MACCE) in this population. METHODS: A total of 3986 patients with ACS were divided into 4 groups based on the quartile (Q) values of peak N-Terminal pro-brain natriuretic peptide (NT-proBNP) measured during hospitalization. The incidence of MACCE was compared among Q1-Q4 groups during follow up. Multivariate Cox regression analysis was performed to identify independent prognostic factors of MACCE. Receiver operating characteristic (ROC) curve was generated to compare the area under the curve (AUC) for MACCE by adding NT-proBNP to the Thrombolysis in Myocardial Infarction (TIMI) risk score. RESULTS: NT-proBNP was significantly positively correlated with peak values of cardiac troponin I (cTnI) (r = 0.418), high-sensitivity C-reactive protein (hs-CRP) (r = 0.397) and left ventricular end-diastolic diameter (LVEDD) (r = 0.075) (P < 0.001). The risks of composite MACCE (5.6%, 9.1%, 13.0%, 20.1%, P < 0.001), all-cause death (1.0%, 2.5%, 4.1%, 8.4%, P < 0.001) and non-fatal myocardial infarction (2.0%, 3.4%, 4.8%, 6.2%, P < 0.001) were significantly higher in the higher Q groups. In multivariate analysis, the Q4 group displayed an independent 2.2-fold increase for MACCE compared to Q1 (HR: 2.16; 95%CI: 1.57-2.99; P < 0.001). Compared with TIMI risk score alone, TIMI+NT-proBNP showed improved AUCs: cardiovascular death (P = 0.0008), and heart failure requiring hospitalization (P = 0.0017). CONCLUSIONS: In patients with ACS with successful PCI and normal LVEF, elevated NT-proBNP was significantly associated with poor clinical outcomes. These results suggest that NT-proBNP is a useful biomarker for prognosis and risk stratification in this population.

Sex-related differences in clinical outcomes and predictive factors in the very elderly patients with ACS undergoing PCI.

Background: As shown in previous studies, there may be sex-related differences in clinical outcomes in patients with acute coronary syndromes (ACS) after percutaneous coronary intervention (PCI). However, the benefits of PCI in very elderly ACS patients and the gender differences were poorly described and understood. We investigated the clinical characteristics and outcomes after PCI by sex stratification, and the predictive factors of major adverse cardiovascular and cerebrovascular events (MACCE) in this very elderly ACS cohort. Methods: A total of 1,676 consecutive ACS patients (50.2% women) aged ≥80 years old between January 2013 and May 2020 were recruited in this study. All patients were divided into four groups according to gender and treatment: male PCI (n = 321) and conservative management groups (n = 513), and female PCI (n = 283) and conservative management groups (n = 559). Clinical and coronary lesion characteristics were compared among four groups, also the clinical outcomes. MACCE and their predictive factors were assessed using Kaplan-Meier curve and Cox regression analysis. Results: PCI procedures were conducted in 604 patients, and 1,072 were conservative management. Men were most likely to present with prior myocardial infarction (MI), peripheral artery disease, and chronic total occlusion (CTO); women had a higher prevalence of hypertension and dyslipidemia. The proportion of men receiving PCI procedures was significantly higher than that of women (38.5 vs. 33.6%, p = 0.038). Compared to conservative management, successful PCI significantly improved composite MACCE in both men (33.9 vs. 18.4%, p < 0.001) and women (27.9 vs. 20.8%, p = 0.026). There were no differences between sex in the improvement of clinical outcomes after PCI. In addition, age, ST-segment elevation myocardial infarction (STEMI), log N-terminal pro-brain natriuretic peptide (NT-proBNP), P2Y12 receptor antagonist, and ß-blocker were independently associated with the incidence of MACCE after PCI tested by the Cox regression model, but not gender (male: hazard ratio (HR) 1.275, 95% confidence interval (CI) 0.853-1.905, p = 0.236). Conclusion: In this very elderly ACS cohort, men presented with more complex clinical conditions, and women were less likely to receive PCI treatment. Both women and men had similar benefits from the PCI procedure in the decrease of MACCE.

[The relationship between left ventricular diastolic function and arterial stiffness in diabetic coronary heart disease].

OBJECTIVES: By measuring left ventricular diastolic function and arterial stiffness, this study aims to probe into the effect of diabetes mellitus (DM) on left ventricular diastolic function and arterial stiffness, and evaluate the correlation between left ventricular diastolic function and arterial stiffness. METHODS: Seventy-six inpatients were enrolled. According to their coronary angiography, OGTT test results and past history of DM, patients were divided into controlled, CHD (coronary heart disease with no DM), and CHD + DM groups. Through invasive hemodynamic monitoring during left ventricular angiography, left ventricular end-diastolic pressure (LVEDP) and tau index were collected. Carotid-femoral pulse wave velocity (c-f PWV), reflected wave augmentation index (AIx@75) and other data reflecting the degree of arterial stiffness were collected bedside with non-invasive means. SPSS 18.0 was used for statistical analysis. RESULTS: No significant difference was found between groups for LVEDP, tau index, and AIx@75. In terms of c-f PMV, The CHD + DM group (8.79 ± 1.59) cm/s differed significantly from the CHD group (7.43 ± 1.42) cm/s and the controlled group (6.83 ± 1.14) cm/s. No correlations were found between c-f PMV and LVEDP or tau index. A positive correlation was found between AIx@75 and tau index. CONCLUSIONS: Compared with the controlled group and CHD patients with no DM, CHD + DM patients show worse arterial stiffness with no difference in ventricular diastolic function. There is a positive correlation between arterial stiffness and diastolic dysfunction.

The Long Terminal Repeats of ERV6 Are Activated in Pre-Implantation Embryos of Cynomolgus Monkey.

Precise gene regulation is critical during embryo development. Long terminal repeat elements (LTRs) of endogenous retroviruses (ERVs) are dynamically expressed in blastocysts of mammalian embryos. However, the expression pattern of LTRs in monkey blastocyst is still unknown. By single-cell RNA-sequencing (seq) data of cynomolgus monkeys, we found that LTRs of several ERV families, including MacERV6, MacERV3, MacERV2, MacERVK1, and MacERVK2, were highly expressed in pre-implantation embryo cells including epiblast (EPI), trophectoderm (TrB), and primitive endoderm (PrE), but were depleted in post-implantation. We knocked down MacERV6-LTR1a in cynomolgus monkeys with a short hairpin RNA (shRNA) strategy to examine the potential function of MacERV6-LTR1a in the early development of monkey embryos. The silence of MacERV6-LTR1a mainly postpones the differentiation of TrB, EPI, and PrE cells in embryos at day 7 compared to control. Moreover, we confirmed MacERV6-LTR1a could recruit Estrogen Related Receptor Beta (ESRRB), which plays an important role in the maintenance of self-renewal and pluripotency of embryonic and trophoblast stem cells through different signaling pathways including FGF and Wnt signaling pathways. In summary, these results suggest that MacERV6-LTR1a is involved in gene regulation of the pre-implantation embryo of the cynomolgus monkeys.

Ginsenoside Rb1 protects cardiomyocytes against CoCl2-induced apoptosis in neonatal rats by inhibiting mitochondria permeability transition pore opening.

AIM: To investigate whether mitochondria permeability transition pore (mPTP) opening was involved in ginsenoside Rb1 (Gs-Rb1) induced anti-hypoxia effects in neonatal rat cardiomyocytes ex vivo. METHODS: Cardiomyocytes were randomly divided into 7 groups: control group, hypoxia group (500 micromol/L CoCl(2)), Gs-Rb1 200 micromol/L group (CoCl(2) intervention+Gs-Rb1), wortmannin (PI3K inhibitor) 0.5 micromol/L group, wortmannin+Gs-Rb1 group, adenine 9-beta-D-arabinofuranoside (Ara A, AMPK inhibitor) 500 micromol/L group, and Ara A and Gs-Rb1 group. Apoptosis rate was determined by using flow cytometry. The opening of the transient mPTP was assessed by using co-loading with calcein AM and CoCl(2) in high conductance mode. Expression of GSK-3beta, cytochrome c, caspase-3 and poly (ADP-ribose) polymerase (PARP) was measured by using Western blotting. DeltaGSK-3beta was defined as the ratio of p-Ser9-GSK-3beta to total GSK-3beta. RESULTS: CoCl(2) significantly stimulated mPTP opening and up-regulated the level of DeltaGSK-3beta. There was a statistically significant positive correlation between apoptosis rate and mPTP opening, between apoptosis rate and DeltaGSK-3beta, and between mPTP opening and DeltaGSK-3beta. Gs-Rb1 significantly inhibited mPTP opening induced by hypoxia (41.3%+/-2.0%, P<0.001) . Gs-Rb1 caused a 77.3%+/-3.2% reduction in the expression of GSK-3beta protein (P<0.001) and a significant increase of 1.182+/-0.007-fold (P=0.0001) in p-Ser9-GSK-3beta compared with control group. Wortmannin and Ara A significantly inhibited the effect of Gs-Rb1 on mPTP opening and DeltaGSK-3beta. Gs-Rb1 significantly decreased expression of cytochrome c (66.1%+/-1.7%, P=0.001), caspase-3 (56.5%+/-2.7%, P=0.001) and cleaved poly ADP-ribose polymerase (PARP) (57.9%+/-1.4%, P=0.001). CONCLUSION: Gs-Rb1 exerted anti-hypoxia effect on neonatal rat cardiomyocytes by inhibiting GSK-3beta-mediated mPTP opening.

Cardiac fibrosis in diabetic rats: regulation and mechanism of activation of the PPARgamma signal pathway.

Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a member of the nuclear hormone receptor superfamily which affects organic fibrosis. The aims of the study were to approach the effects of activation of the PPARgamma signal pathway on cardiac fibrosis in diabetic rats, and also the effects on cardiac remodeling and function. Type 1 diabetic models were used in the study. All the animals were divided into 3 groups: I: control group; II: diabetic group; III: diabetes+Pioglitazone (Piog, a PPARgamma ligand) administration group. After 14 weeks of feeding, general condition, fibrosis indices, echocardiography and interventricular pressures parameters were detected. At the 14th week, compared with group I, the hydroxyproline concentration in group II significantly increased, and CO I and III distribution was more obvious by sirius red staining. Reduction of LVSP (left ventricular systolic pressure) and increase of LVEDP (left ventricular end-diastolic pressure) were also significant in group II. But these situations were changed by the administration of Piog in group III. Furthermore, results of RT-PCR and immunohistochemistry showed that Piog administration reduced angiotensin II type 1 receptor (AT1-R) expression in diabetic models. Hence, activation of the PPARgamma signal pathway could repress cardiac fibrosis in diabetic rats, and partly improve cardiac remodeling and function by down-regulating activity of RAS at the receptor level.

Repeated Herbal Acupoint Sticking Relieved the Recurrence of Allergic Asthma by Regulating the Th1/Th2 Cell Balance in the Peripheral Blood.

Allergic asthma is an inflammatory disease involving the Th1/Th2 cell imbalance in the peripheral blood. Repeated herbal acupoint sticking (RHAS) has been used for hundreds of years in China to relieve the recurrence of allergic asthma, and it is still practiced today. Thus, we explored the effect on allergic asthma relapse and the underlying immunoregulatory mechanism in this study. Here, we enrolled 50 allergic asthma participants, and 38 of them completed the treatment and follow-up (the allergic asthma group). In addition, 13 healthy participants (the control group) were enrolled. The recurrence number of allergic asthma participants and asthma control test (ACT) were used to evaluate the effect of treatment on relieving allergic asthma recurrence. Flow cytometry was performed to analyze the levels of Th1 and Th2 cells in the peripheral blood. The serum levels of IgE, IFN-γ, and IL-4 were detected by ELISA. (1) In the allergic asthma group, compared to before the first treatment, the recurrence number of allergic asthma participants decreased and the ACT score increased at end of the last treatment, 18 and 30 weeks of the trial (P < 0.05). At 18 and 30 weeks of the trial, the recurrence number of allergic asthma participants was less and the ACT score was higher than the ones from the same period last year in the allergic asthma group (P < 0.05). Compared to before the first treatment, the percentage of Th1 cell did not change significantly, the percentage of Th2 cell decreased, and the Th1/Th2 cell ratio increased in the allergic asthma group by the end of the last treatment (P < 0.05). Meanwhile, the release of IgE and IL-4 reduced (P < 0.05), and the release of IFN-γ did not significantly change in the allergic asthma group. (2) Compared with the control group, the serum levels of IgE and IL-4 and the percentage of Th2 cell were higher, and the Th1/Th2 cell ratio was lower in the allergic asthma group (P < 0.05). There was no significant difference between Th1 cell and IFN-γ before the first treatment. (3) Compared with the control group, the IgE levels and the percentage of Th2 cell were higher in the allergic asthma group (P < 0.01). Simultaneously, there was no significant difference between Th1 cell, the Th1/Th2 cell ratio, and the serum levels of IFN-γ and IL-4 by the end of the last treatment. The data suggested that RHAS reduced the amount of Th2 cell and elevated the Th1/Th2 cell ratio, thereby alleviating the inflammatory responses in the allergic asthma participants.

Anti-hypoxic effect of ginsenoside Rbl on neonatal rat cardiomyocytes is mediated through the specific activation of glucose transporter-4 ex vivo.

AIM: The aim of this study was to investigate whether Gs-Rbl relieves the CoCl(2)-induced apoptosis of hypoxic neonatal rat cardiomyocytes and in which the role of glucose transporter-4 (GLUT-4). METHODS: Gs-Rbl (0, 10, 50, 100, 200, 400, and 500 micromol/L), adenine 9-beta-D-arabinofuranoside (ara A, 500 micromol/L; AMPK inhibitor) and wortmannin (0.5 micromol/L; PI3K inhibitor) only in combination with 200 micromol/L Gs-Rbl were administered in hypoxic cardiomyocytes, which were induced by 500 micromol/L CoCl(2) for 12 h. Then, the apoptotic rate (AR), 2-[(3)H]-deoxy-D-glucose (2-[(3)H]-DG) uptake, and the expression of GLUT-4 (including in plasma membrane, PM), phospho-AMPKalpha (Thr172), AMPKalpha and Akt in cells were assayed. RESULTS: Compared with simple hypoxia (0 micromol/L Gs-Rbl), Gs-Rb1 greater than 10 micromol/L significantly decreased the apoptotic rate (P<0.01) and significantly increased 2-[(3)H]-DG uptake (P<0.01), GLUT-4 content in cells and PM (P<0.01), AMPK activity (P<0.01) and Akt (P<0.01) levels in a dose-dependent manner. AMPK activity was completely suppressed by ara-A, just as Akt was suppressed by wortmannin. The AR, glucose uptake and GLUT-4 levels in cells and PM were partly down-regulated by ara-A or wortmannin. CONCLUSION: Gs-Rb1 may protect neonatal rat cardiomyocytes from apoptosis induced by CoCl(2). The anti-apoptotic effect of Gs-Rb1 may occur by improving glucose uptake, in which GLUT-4 translocation and expression played a key role. Both the AMPK and the PI3K/Akt pathways may take part in the anti-hypoxic efficacy of Gs-Rb1.

Ginsenoside Rb1 Ameliorates Autophagy of Hypoxia Cardiomyocytes from Neonatal Rats via AMP-Activated Protein Kinase Pathway.

OBJECTIVE: To investigate whether ginsenoside-Rb1 (Gs-Rb1) improves the CoCl-induced autophagy of cardiomyocytes via upregulation of adenosine 5'-monophosphate-activated protein kinase (AMPK) pathway. METHODS: Ventricles from 1- to 3-day-old Wistar rats were sequentially digested, separated and incubated in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum for 3 days followed by synchronization. Neonatal rat cardiomyocytes were randomly divided into 7 groups: control group (normal level oxygen), hypoxia group (500 µmol/L CoCl2), Gs-Rb1 group (200 µmol/L Gs-Rb1 + 500 µmol/L CoCl2), Ara A group (500 µmol/L Ara A + 500 µmol/L CoCl2), Ara A+ Gs-Rb1 group (500 µmol/L Ara A + 200 µmol/L Gs-Rb1 + 500 µmol/L CoCl2), AICAR group [1 mmol/L 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) + 500 µmol/L CoCl2], and AICAR+Gs-Rb1 group (1 mmol/L AICAR + 200 µmol/L Gs-Rb1 + 500 µmol/L CoCl2). Cells were treated for 12 h and cell viability was determined by methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay and cardiac troponin I (cTnI) levels were detected by enzyme-linked immunosorbent assay (ELISA). AMPK activity was assessed by 2',7'-dichlorofluorescein diacetate (DCFH-DA) ELISA assay. The protein expressions of Atg4B, Atg5, Atg6, Atg7, microtubule-associated protein 1A/1B-light chain 3 (LC3), P62, and active-cathepsin B were measured by Western blot. RESULTS: Gs-Rb1 significantly improved the cell viability of hypoxia cardiomyocytes (P<0.01). However, the viability of hypoxia-treated cardiomyocytes was significantly inhibited by Ara A (P<0.01). Gs-Rb1 increased the AMPK activity of hypoxia-treated cardiomyocytes. The AMPK activity of hypoxia-treated cadiomyocytes was inhibited by Ara A (P<0.01) and was not affected by AICAR =0.983). Gs-Rb1 up-regulated Atg4B, Atg5, Beclin-1, Atg7, LC3B II, the LC3B II/I ratio and cathepsin B activity of hypoxia cardiomyocytes (P<0.05), each of these protein levels was significantly enhanced by Ara A (all P<0.01), but was not affected by AICAR (all P>0.05). Gs-Rb1 significantly down-regulated P62 levels of hypoxic cardiomyocytes (P<0.05). The P62 levels of hypoxic cardiomyocytes were inhibited by Ara A (P<0.05) and were not affected by AICAR (P=0.871). CONCLUSION: Gs-Rb1 may improve the viability of hypoxia cardiomyocytes by ameliorating cell autophagy via the upregulation of AMPK pathway.

Down-regulation of the expression of angiotensin II type 1 receptor in neonatal rat cardiac fibroblast by activation of PPARgamma signal pathway.

Peroxisome proliferator-activated receptor-gamma (PPARgamma) is one of the hormone nuclear receptors. Recent data have shown that activation of PPARgamma signal pathway has many positive effects on cardiovascular system. The goals of this study were to determine whether PPARgamma activator affects cardiac fibrosis and the possible mechanisms. Cardiac fibroblasts (CFs) of SD neonate rats were used in the study. Cells were divided into 4 groups: I--control group; II--pioglitazone group (Piog--PPARgamma agonist); III--angiotensin II (Ang II) group; and IV--Piog + Ang II group (Piog plus angiotensin II). mRNA and protein expression of collagen type I, III and angiotensin II type 1 receptor (AT1-R) were tested by reverse transcription--polymerase chain reaction and Western blotting. With the inhibition of actinomycin D, we investigated the impacts of Piog on the stability of AT1-RmRNA. Compared with group I, the mRNA and protein expression of collagen type I, III and AT1-R were up-regulated in group III (P < 0.05). However with the effects of Piog in group IV, the expressions mentioned above were attenuated significantly (P < 0.05). With the effects of actinomycin D, AT1-RmRNA was reduced at the same degree in control and Piog groups at the same time points. These results indicated that treatment with Piog can attenuate Ang II-induced collagen synthesis in CFs through down-regulation of the AT1-R expression. With the intervention of actinomycin D, we suggested that PPARgamma agonist didn't affect the stability of AT1-RmRNA.

Genetic characterization of plasmid-mediated quinolone resistance gene qnrS2 in Pseudoalteromonas and Shewanella isolates from seawater.

Three qnrS2-containing isolates of Pseudoalteromonas and Shewanella were collected from the seawater samples of Qingdao in China during 2014. They displayed resistance to ampicillin, ciprofloxacin, kanamycin, nalidixic acid and sulfamethoxazole. The qnrS2 genes were identified in the chromosomes of Pseudoalteromonas strains E8 and S16, and in a 140-kb plasmid in Shewanella strain S14, respectively. In addition, two copies of qnrS2 were identified in the strain E8. Sequence analyses revealed that there was an identical DNA segment located in the downstream of qnrS2 in strain S14 and E8, coding for a TetR transcriptional regulator, two putative integrases and a hypothetical protein. However, different genetic structures were identified in the upstream sequences: the terB gene associated with tellurite resistance in the strain S14, and a putative integron with dfrA6 and aadA13 gene cassettes or the Tn7-related gene complex tnsABC in the strain E8. In Pseudoalteromonas strain S16, qnrS2 was bracketed by the endonuclease I and III genes, and the electron transport complex rsxCDGE was located in the upstream sequences. This is the first report of two copies of the qnrS2 gene existing in one bacterial chromosome, and also the first identification of qnrS2 in Shewanella.

Apelin-APJ effects of ginsenoside-Rb1 depending on hypoxia-induced factor 1α in hypoxia neonatal cardiomyocytes.

OBJECTIVE: To investigate whether ginsenoside-Rb1 (Gs-Rb1) inhibits the apoptosis of hypoxia cardiomyocytes by up-regulating apelin-APJ system and whether the system is affected by hypoxia-induced factor 1α (Hif-1α). METHODS: Neonatal rat cardiomyocytes were randomly divided into 6 groups: a control group, a simple CoCl group, a simple Gs-Rb1 group, a CoCl and Gs-Rb1 hypoxia group, a CoCl and 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1) group, a CoCl and YC-1 group and a Gs-Rb1 group, in which YC-1 inhibits the synthesis and accelerates the degradation of Hif-1a. The concentration of CoCl, Gs-Rb1 and YC-1 was 500 µmol/L, 200 µmol/L and 5 µmol/L, respectively; the apoptosis ratio was analyzed with a flow cytometer; and apelin, APJ and Hif-1α were assayed with immunocytochemistry, Western blot assays and reverse transcription polymerase chain reaction (RT-PCR). RESULTS: (1) The anti-apoptosis effect of Gs-Rb1 on hypoxia cardiomyocytes was significantly inhibited by YC-1; (2) Hypoxia significantly up-graded the expression of mRNA and protein of apelin; this effect was further reinforced by Gs-Rb1 and significantly inhibited by YC-1; (3) Gs-Rb1 further strengthened the expression of APJ mRNA and APJ proteins once hypoxia occurred, which was significantly inhibited by YC-1; (4) Gs-Rb1 significantly increased the expression of Hif-1α, which was completely abolished by YC-1; (5) There was a negative relationship between AR and apelin (or APJ, including mRNA and protein), a positive correlation between apelin (or APJ) protein and Hif-1a protein, in hypoxia cardiomyocytes. CONCLUSION: The apelin-APJ system plays an important role in the anti-apoptosis effect of Gs-Rb1 on hypoxia neonatal cardiomyocytes, which was partly adjusted by Hif-1α.

Admission glucose level and in-hospital outcomes in diabetic and non-diabetic patients with ST-elevation acute myocardial infarction.

BACKGROUND: Hyperglycemia on admission is a predictor of an unfavorable prognosis in patients with ST-elevation Acute Myocardial Infarction (AMI). Data concerning associations between an elevated glucose level on admission and other in-hospital complications are still limited. METHODS: A total of 1,137 AMI patients with complete admission blood glucose level (ABGL) analysis were identified and stratified according to ABGL. RESULTS: A total of 16.1% patients had admission glucose level <5 mmol/L, 36.1% <7 mmol/L, 20.2% <9 mmol/L, 9.9% <11 mmol/L and 17.7% ≥11 mmol/L. Compared with the euglycemia group, both the hypo- and hyperglycemia groups were associated with higher in-hospital mortality. In-hospital mortality of diabetic patients with hypoglycemia (12.2%) was higher than that of diabetic patients with either euglycemia or mild hyperglycemia (11.1%, or 10.7% relatively). The same results were seen in non-diabetic patients. In the logistic regression analysis, admission glucose and cardiac function of Killip grade were the independent predictors of in-hospital death for patients with AMI. CONCLUSION: Elevated admission glucose levels are associated with an increased risk of life-threatening complications in diabetic and non-diabetic AMI patients. Compared with the euglycemia group, hypoglycemia was associated with a higher trend of in-hospital mortality.

Predictive value of serum uric acid on cardiovascular disease and all-cause mortality in urban Chinese patients.

BACKGROUND: The association between increased serum uric acid (SUA) levels and cardiovascular risk has been debated for decades. Several large studies have provided conflicting results regarding the clinical significance of elevated SUA levels in cardiovascular disease (CVD) or cerebrovascular disease. The aim of this study was to investigate the relationship between SUA and CVD and all-cause mortality and their potential diagnostic value. METHODS: A total of 3570 in-patients ranging in age from 56 to 95 years (mean (67.36 +/- 11.36) years) were selected from 20 hospitals in Beijing and Shanghai. A carefully designed questionnaire was used to gather baseline data of each patient. All patients were divided into two main groups according to their SUA levels: high SUA and normal SUA groups. Serum indices and other important parameters were measured. RESULTS: Compared with normal SUA group, high SUA group had significant difference in systolic blood pressure (SBP), total cholesterol (TC), triglyceride (TG), high density lipoprotein cholesterol (HDL-C), body mass index (BMI), and age (P < 0.05 or P < 0.01). High SUA prevailed in female and patients with history of essential hypertension, while history of smoking and diabetes showed no significant difference between two groups. All-cause and CVD mortality occurred more frequently in high SUA group than in normal SUA group. In the accumulative survival analysis, high SUA group had lower survival rate than normal SUA group both in CVD and all-cause mortality. COX regression analysis indicated that the history of smoking, age and high SUA were independent risk factors for the development of CVD. CONCLUSIONS: These preliminary observations suggest that patients with high SUA levels would face higher risk of mortality. SUA measurement may be applied as a routine predictor for clinical assessment.

Relation of Hemoglobin A1c to myocardial acoustic densitometry and left ventricular diastolic function in patients with Type 2 diabetes mellitus and without evident heart disease.

AIM: The aim of the study was to detect the echocardiographic sensitive indexes for prediction of the subclinical cardiac dysfunction and to evaluate the relation between Hemoglobin A1c (HbA1c) and myocardial acoustic densitometry as well as cardiac function. METHODS: Fifty DM2 patients (48.5+/-8.6 years) without evident heart disease and 50 age- and sex-matched normal controls (47.6+/-8.8 years) were enrolled. Conventional echocardiography, tissue Doppler imaging and acoustic densitometry of both groups were measured. HbA1c of DM2 patients were determined. RESULTS: There were no significant differences in systolic indexes between the two groups. Mean LV myocardial early diastolic velocity Em and Em/Am were lower in DM group than control group. Mean LV myocardial late diastolic velocity Am and E/Em were higher in DM group than control group. IVS-IBS% and LVPW-IBS% were higher in DM group than control group. IVS-CVIB and LVPW-CVIB were lower in DM group than control group. HbA1c was negatively correlated with E/A (gamma=-0.310, P<0.05), Em (gamma=-0.409, P<0.01), Em/Am (gamma=-0.380, P<0.01) and positively correlated with E/Em (gamma=0.488, P<0.01). Significant positive correlation was present between HbA1c and IVS-IBS% (gamma=0.679, P<0.01), LVPW-IBS% (gamma=0.666, P<0.01). CONCLUSIONS: The diastolic dysfunction and abnormal myocardial acoustic densitometry exist before the systolic function damage in DM2 patients. Tissue Doppler imaging and ultrasonic integrated backscatter can be used as sensitive means for early assessing myocardial histological changes in DM2 patients. HbA1c is related with both diastolic dysfunction and acoustic densitometry.